The renal thick ascending limb contributes to the maintenance of acid-base homeostasis by reabsorbing both bicarbonate and ammonium. Our long-term goal is to understand the cellular, biochemical, and molecular mechanisms by which these transport processes are regulated. The central hypothesis of this proposal is that the control of HCO3 absorption in the medullary thick ascending limb (MTAL) depends on the regulation of both apical and basolateral Na+/H+ exchangers. We have developed evidence of a novel mechanism for regulation of HCO3 absorption in the MTAL, in which the activity of the apical Na+/H+ exchanger, which is directly involved in HCO3 absorption, is regulated by the activity of the basolateral Na+/H+exchanger. The current studies are directed toward understanding the mechanism whereby basolateral Na+/H+ exchange regulates apical Na+/H+ exchange, and identifying physiological factors and signaling pathways that regulate cross-talk between the exchangers. Preliminary studies show that the cross-talk mechanism is genetically eliminated in MTAL from NHE1 knockout mice, which will be studied to define the role of basolateral Na+/H+ exchange in regulating apical Na+/H+exchange and HCO3 absorption. The Specific Aims include: 1) determine the role of the actin cytoskeleton in mediating regulation of the apical Na+/H+ exchanger by the basolateral Na+/H+ exchanger, 2) examine chronic regulation of the interaction between basolateral and apical Na+/H + exchangers by changes in dietary Na+ intake, and 3) examine acute regulation of cross-talk between the exchangers by growth factors and aldosterone via extracellular signal-regulated kinase- and phosphatidylinositol 3-kinase-dependent pathways. Biochemical analyses and fluorescence microscopy will be integrated with ion transport measurements in MTALs microdissected from rats and gene-targeted mice to provide a detailed understanding of the cross-talk mechanism and its role in HCO3 absorption. These studies will identify novel mechanisms for the regulation of Na+/H + exchange activity and acid secretion in renal tubules and will investigate a new biological process involving cross-talk between Na+/H+ exchangers in basolateral and apical membranes of epithelial cells. The results will enhance our understanding of the role of the MTAL in the physiological and pathophysiological control of acid-base balance. In addition, they will be the first to identify a function for NHE1 in transepithelial transport in the kidney and to define a role for this basolateral exchanger in the acute and chronic regulation of HCO3 absorption.